Vitamin D is an integral component of the mechanism by which higher organisms regulate calcium, phosphorus, and skeletal homeostasis. Recently broader roles for vitamin D in most tissues of the body have been suggested. The active metabolite of vitamin D, 1,25(OH)2-vitamin D, acts largely, if not exclusively by binding to an intracellular protein, the vitamin D receptor. This receptor, activated by the binding of 1,25(OH)2-vitamin D, then modulates the expression of specific genes. A detailed understanding of receptor function will require in vitro analysis of the interaction of purified receptor with 1,25(OH)2vitamin D and with specific target genes, as well as the analysis of the behavior of normal and mutant receptor genes in cultured cells. The scarcity of the protein has hampered efforts to purify enough receptor for detailed in vitro characterization. This project will involve purification of nucleic acids encoding the chick, rat, and human vitamin D receptors. The amino acid sequences of the proteins will be deduced from the DNA sequences, and expresson vectors will be used to synthesize large amounts of receptor protein in bacteria and in mammalian cells. The functional importance of discrete portions of the receptor will be defined by analyzing the sequences of the abnormal receptors carried by patients with vitamin D-dependent rickets, type II. Finally, we shall be able to introduce normal and mutant receptor-encoding sequences into cultured cells lacking receptors and thereby study the funciton of normal and abnormal receptors in intact cells. These studies will lead to a detailed understanding of how the vitamin D receptor interacts with 1,25(OH)2-vitamin D and subsequently activates the expression of specific genes. This work will thus yield a greater understanding of the mechanism of vitamin D action in ensuring calcium homeostasis in health and disease.